The major cellular phosphatase Protein Phosphatase 2A (PP2A) is regulated by an unconventional but conserved methylation modification on the very carboxy terminus of its catalytic subunit. However, the functional role of this methylation remains incompletely understood. In the current funding period, it was shown that this methylation modification on PP2A is responsive to intracellular methionine and SAM levels. As such, the amino acid methionine and its downstream metabolite SAM are able to promote the methylation of PP2A and thereby influence the activity of the phosphatase against particular substrates in cellular signaling and growth control. This renewal application proposes to comprehensively investigate the in vivo functions of the methylation of PP2A using a combined genetic, chemical, and proteomic approach. It is notable that this modification on PP2A enables it to function as an amino acid-responsive, metabolically-regulated phosphatase which could impact the phosphorylation status of many important signaling proteins in tune with the metabolic state of the cell.